Mesenchymal stem cells were first found in the mouse spinal cord cells, which were in the form of fibroblasts (Friedenstein et al., Cell Tissue Kinet, 20, 263-272, 1987). Mesenchymal stem cells are classified into several mesodermal series and these cells can be differentiated into various cells, that is mesenchymal stem cells are characterized by their diversity in differentiation into osteoblasts (Friedenstein et al., Cell Tissue Kinet, 20, 263-272, 1987; Abdallah et al., Bone, 39, 181-188, 2006), adipocytes (Fink et al., Stem Cells, 22, 1346-1355, 2004; Abdallah et al., Bone, 39, 181-188, 2006), endothelial cells (Kassem et al., Basic Clin Pharmacol Toxicol 95, 209-14, 2004) and neuronal cells (Woodbury et al., J Neurosci Res, 61, 364-370, 2000; Woodbury et al., J Neurosci Res, 69, 908-917, 2002; Krabbe et al., APMIS, 113, 831-844, 2005).
Through clinical trials, understanding of mesenchymal stem cells has been widened and studies on the efficiency and stability of gene therapy based on transplantation have been actively undergoing (Zhao et al., J Neurol Sci, 233, 87-91, 2005). Recently, it has been reported that bone marrow-derived mesenchymal stem cells were applied to induce regeneration of bone, myocardium and nerves (Jiang et al., Zhonghua Er Bi Yan Hou Ke Za Zhi, 37, 137-139, 2002; Kassem et al., Basic Clin Pharmacol Toxicol, 95, 209-214, 2004; Kemp et al., Leuk Lymphoma, 46, 1531-1544, 2005; Biossy et al., Cancer Res 65, 9943-9952, 2005; Krabbe et al., APMIS, 113, 831-844, 2005). In particular, this application seems to have great advantage for the treatment of various neurodegenerative diseases because the transplantation of stem cells into the wounded area not only opens the possibility of nerve replacement but also plays an important role in maintaining existing cells (Maragakis et al., Glia, 50, 145-159, 2005).
Chronic pathological phenomena occurring after short-term damage on spinal nerves have been studied and in fact characteristics or symptoms of in vivo spinal cord injury models have been reported (Krassioukov et al., J Neurotrauma, 19, 1521-1529, 2002; Lee et al., Orthop Clin North Am, 33, 311-315, 2002; Casha et al., Exp Neurol, 196, 390-400, 2005; Goldman, Nat Biotechnol, 23, 862-871, 2005). From the observation on the area neighboring damaged spinal cord was confirmed that direct damage or tissue destruction was reduced as farther from the damaged area (Buss & Schwab, Glia, 42, 424-432, 2003) and spinal cord injury was associated with the loss of neurons and glia (Goldman, Nat Biotechnol, 23, 862-871, 2005). Once the spinal cord is damaged, changes in expression of various genes and proteins are observed, which finally cause demyelination, according to recent reports (Di Giovanni et al., Proc Natl Acad Sci USA., 102, 8333-8338, 2005; Kang et al., Proteomics, 6, 2797-2812, 2006). In vivo spinal cord injury model makes the interpretation of the results from experiments difficult because of the complexity of the in vivo system. Thus, in vitro model has been preferred, since it enables the regulation from the outside of a cell and the repetition of the experiment. And it costs less. However, such in vitro model dose not have any biological activity.
Organotypic slice cultures have been used for various injury models for the study of ischemia and cytotoxicity (Krassioukov et al., J Neurotrauma, 19, 1521-1529, 2002). Spinal cord culture exhibits distinguishable dorsal horn and ventral horn and is in excellent shape as a whole, suggesting that it is easy to observe neuronal distribution (Ulich et al., J Neurophysiol., 72, 861-871, 1994; Takuma et al., Neuroscience, 109, 359-370, 2002; Hilton et al., Brain Res Brain Res Rev, 46, 191-203, 2004).
Demyelination of axon without structural damage and the disorder of saltatory conduction might be the cause of functional defect by spinal cord injury (Nicot, Brain 126, 398-412, 2003). Lysolecithin is a lipid containing detergent-like protein and has a similar activity with a membrane soluble agent exhibiting myelinated cell-specific toxicity to cause demyelination (Franklin et al., J Neurosci Res 58, 207-213, 1999; Arnett et al., Science, 306, 2111-2115, 2004). It has been known that lysolecithin causes demyelination when it is inserted in myelinated fiber in tissues such as the spinal cord and thus causes spinal cord injury in vivo. In the meantime, the loss of oligodendrocyte, a cause of various adult demyelination-associated diseases including multiple sclerosis, has been proved to be associated with demyelination of spinal cord. When the precursor cells of human oligodendrocytes were transplanted in the adult mouse brain damaged by lysolecithin, the precursor cells were rapidly differentiated into oligodendrocytes-like cells and induced remyelination of the demyelinated axon although the efficiency was lower than true myelinating cells (Goldman, Nat Biotechnol, 23, 862-871, 2005). These results suggest that axons demyelinated by lysolecithin can be remyelinated by the transplantation of mesenchymal stem cells.
The present inventors confirmed that neuronal regeneration of damaged axons in spinal cord slices could be induced after transplantation of bone marrow-derived mesenchymal stem cells into those slices which had previously been demyelinated by a toxic compound. Furthermore, the present inventors completed this invention by confirming that directional neuronal regeneration could be effectively applied as a novel treatment method for neurodegenerative diseases such as spinal cord injury.